Poly(ethylene glycol) (PEG) hydrogel has been successfully utilized as a cell-encapsulation material to replicate the microenvironments of various tissues. However there has been limited development of new networks with improved permeability to support metabolic activities of encapsulated cells, while maintaining patternability to recapitulate key aspects of tissue architecture.
Engineered tissues are expected to overcome shortcomings associated with traditional tissue replacement therapies and provide a platform to study various biological assays in vitro. Several types of cells cultured as two-dimensional monolayers are known to de-differentiate and lose their original functions due to disruption of their normal microenvironments. Thus, it has been a major challenge in the field of tissue engineering to provide structural and functional support equivalent to native tissues. Use of hydrogels as a scaffold material for engineered tissues has been of increasing interest due to their structural similarity to the extracellular components in the body. Photopolymerizable polyethylene glycol) (PEG) is one of the most extensively utilized hydrogels because its network structure can be easily modified to mimic critical aspects of the original microenvironments. Various approaches to design PEG networks for improved phenotype stability of encapsulated cells have involved conjugating other biologically active factors to the polymer network and incorporating degradable linkages. Another advantage of PEG is that use of photolithography or microfluidic processing allows fabrication of microarchitectures that potentially recapitulate key aspects of tissue architecture to guide cells' behavior with respect to morphology, cytoskeletal structure, and functionality (see Khademhosseini et al., Microscale technologies for tissue engineering and biology. PNAS 2006, 103, (8), 2480-2487).
A three-dimensional matrix of an engineered tissue needs to provide sufficient permeability to support metabolic activities, provide for unimpeded transport of large macromolecules, and permit multiple cell type interactions that are normally present in most original tissues (Sachlos et al., Making tissue engineering scaffolds work. Review: the application of solid freeform fabrication technology to the production of tissue engineering scaffolds. Eur Cell Mater 2003, 5, 29-39). Even though the matrix diffusion condition could be gradually improved by promoting angiogenesis or conjugating degradable linkages, it is required to allow sufficient diffusion of substances indispensable to cell survival right after cells are encapsulated. In addition, because many scaffolds for tissue engineering initially need to fill a space and provide a framework for the replaced tissue, the mechanical properties of the material are important. However, it is a challenge for hydrogel-based scaffolds to allow sufficient permeability, while maintaining mechanical properties sufficient for generating and sustaining reconstructed tissue architectures.
In many polymeric materials, improved permeability of networks has been obtained by increasing the distance between consecutive crosslinks. However, the traditional approach using higher molecular weight macromonomers to increase the distance between crosslink junctions also results in increased viscosity of the liquid phase prepolymer. This retards flow of the prepolymer solution through micro-scale architectures and also leads to decreased mechanical strength of the cured hydrogel. There have been attempts to address the relationship between the traditional polymeric parameters and the behavior of encapsulated cells in PEG networks. However, because there has been no clear explanation of the effect of cell encapsulation protocols on the PEG network structure, development of new networks has been limited.
The present invention addresses the design of networks for improved viability of the encapsulated cells. Network defects by are augmented to increase permeability. This new design has led to improved viability and function of encapsulated cells and reliable control over spatial distribution of incorporated cells at the micron-scale within cell-encapsulated three-dimensional PEG matrices.